A simple method for removing low-density granulocytes to purify T lymphocytes from peripheral blood mononuclear cells

Zhang, Sigong; Song, Yuxin; Shu, Xi; Shen, Hongfang; Yang, Hanbo; Duo, Rui-xue; Wang, Guochun

doi:10.1631/jzus.b1600064

Cited by 6 publications

(3 citation statements)

References 21 publications

(37 reference statements)

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“…These results confirm that LDGs indeed can inhibit the secretion of IFN-γ in T cells through some mechanism. However, a previous study by Zhang et al had reported an opposite impact of LDG removal on T-SPOT assay ( Zhang et al, 2017 ). In this study, LDGs are removed by PBMCs adherence method and whole blood sedimentation method.…”

Section: Discussionmentioning

confidence: 94%

Low-Density Granulocytes Affect T-SPOT.TB Assay by Inhibiting the Production of Interferon-γ in T Cells via PD-L1/PD-1 Pathway

Rao

Peng

et al. 2021

Front. Microbiol.

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BackgroundT-SPOT TB (T-SPOT) assay is widely used for detection of Mycobacterium tuberculosis infection that is based on the detection of M. tuberculosis-specific interferon-γ-secreting T cells (ISCs) in peripheral blood mononuclear cells (PBMCs). Recently, high frequencies of low-density granulocytes (LDGs) were found in the PBMCs of tuberculosis patients. Whether these LDGs affect the detection of T-SPOT has not been investigated. The impact of LDGs on T-SPOT assay and related mechanism were investigated in this study.MethodsThe correlations between the frequencies of LDGs and the results of T-SPOT were analyzed. T-SPOT with LDG-removed PBMCs and PBMCs with exogenous addition of LDGs were performed. The possible mechanism was explored by detecting the levels of negative immune regulatory molecules on LDGs. The impact of programmed death ligand 1 (PD-L1) on T-SPOT was evaluated and confirmed by function blocking with neutralizing antibody.ResultsThe positive rates of T-SPOT and ISCs in tuberculosis patients with low LDGs frequency (n = 22) were significantly higher than those with high LDGs frequency (n = 39). Removal or exogenous addition of LDGs significantly increased or decreased the ISCs and the positive rate of T-SPOT. The frequencies of interferon-γ-producing T cells were negatively correlated with the frequencies of LDGs. The expression of PD-L1 was significantly elevated on LDGs. Pretreatment of LDGs with anti-PD-L1 antibody significantly counteracted the impact of LDGs on T-SPOT. Treatment of PBMCs with anti-PD-L1 antibody resulted in comparable ISCs with that of LDG removal.ConclusionLDGs can inhibit the production of interferon-γ in T cells and decrease the positive rated of T-SPOT assay via highly expressed PD-L1.

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Section: Discussionmentioning

confidence: 94%

Low-Density Granulocytes Affect T-SPOT.TB Assay by Inhibiting the Production of Interferon-γ in T Cells via PD-L1/PD-1 Pathway

Rao

Peng

et al. 2021

Front. Microbiol.

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“…Similarly, to normal-density granulocytes (NDG), LDG neutrophils strongly express LDG-specific surface markers (CD10, CD15 and CD16), but, in contrast to NDG, their nucleus is immature [90,91]. LDGs exhibit features of reactive cells.…”

Section: Low-density Granulocytes (Ldg) and Neutrophil Extracellular Traps (Nets) In Aihmentioning

confidence: 99%

Etiopathogenesis and Diagnostic Strategies in Autoimmune Hepatitis

et al. 2021

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Autoimmune hepatitis (AIH) is a chronic liver disease with the incidence of 10 to 17 per 100,000 people in Europe. It affects people of any age, but most often occurs in the 40–60 age group. The clinical picture is varied, from asymptomatic to severe acute hepatitis or liver failure. The disease onset is probably associated with the impaired function of T lymphocytes, the development of molecular mimicry, intestinal dysbiosis, or infiltration with low density neutrophils, which, alongside autoantibodies (i.e., ANA, ASMA), implicate the formation of neutrophil extracellular traps (NETs), as a component of the disease process, and mediate the inappropriate immune response. AIH is characterized with an increased activity of aminotransferases, elevated concentration of serum immunoglobulin G, the presence of circulating autoantibodies and liver inflammation. The result of the histological examination of the liver and the presence of autoantibodies, although not pathognomonic, still remain a distinguishing feature. The diagnosis of AIH determines lifelong treatment in most patients. The treatment is implemented to prevent the development of cirrhosis and end-stage liver failure. This work focuses mainly on the etiopathogenesis and diagnosis of AIH.

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“…The number of CD4 + T lymphocytes was assessed using a FACSCalibur type flow cytometer and a tricolor fluorescent antibody labeling kit (BD, USA), as described by Yao et al (2017) and Zhang SG et al (2017). According to the reagent specification, each sample was processed in a TruCONT tube dedicated to flow cytometry.…”

Section: Detection Of Cd4 + T Lymphocytes By Flow Cytometrymentioning

confidence: 99%

Changes in peripheral blood inflammatory factors (TNF-α and IL-6) and intestinal flora in AIDS and HIV-positive individuals

Zhang

et al. 2019

J. Zhejiang Univ. Sci. B

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Objective: In this study, we investigated the changes in peripheral blood inflammatory factors and intestinal flora in acquired immune deficiency syndrome (AIDS) and human immunodeficiency virus (HIV)-positive individuals (AIDS/HIV patients), and explored the relationships among intestinal flora, peripheral blood inflammatory factors, and CD4 + T lymphocytes. Methods: Thirty blood and stool samples from an AIDS group and a control group were collected. The levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were determined by enzyme-linked immunosorbent assay (ELISA), and the number of CD4 + T lymphocytes by a FACSCount automated instrument. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the messenger RNA (mRNA) levels of Bifidobacterium, Lactobacillus, Escherichia coli, Enterococcus faecalis, and Enterococcus faecium. Correlations among intestinal flora, inflammatory factor levels, and CD4 + T lymphocyte values were evaluated using the Spearman correlation coefficient. Results: The levels of TNF-α and IL-6 in the AIDS group were higher than those in the control group, while the number of CD4 + T lymphocytes was lower. The amounts of Bifidobacterium and Lactobacillus in the AIDS group were significantly lower than those in control group, while the amounts of E. coli, E. faecalis, and E. faecium were much higher. The amounts of Bifidobacterium and Lactobacillus were negatively correlated with the content of TNF-α and IL-6 and the CD4 + T lymphocyte count, while those correlations were reversed for E. coli, E. faecalis, and E. faecium. Conclusions: The intestinal microbiota of AIDS/HIV patients were disordered, and there was a correlation between the amount of intestinal flora and the number of CD4 + T lymphocytes and the levels of TNF-α and IL-6.

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A simple method for removing low-density granulocytes to purify T lymphocytes from peripheral blood mononuclear cells

Cited by 6 publications

References 21 publications

Low-Density Granulocytes Affect T-SPOT.TB Assay by Inhibiting the Production of Interferon-γ in T Cells via PD-L1/PD-1 Pathway

Low-Density Granulocytes Affect T-SPOT.TB Assay by Inhibiting the Production of Interferon-γ in T Cells via PD-L1/PD-1 Pathway

Etiopathogenesis and Diagnostic Strategies in Autoimmune Hepatitis

Changes in peripheral blood inflammatory factors (TNF-α and IL-6) and intestinal flora in AIDS and HIV-positive individuals

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